Breast cancer is affected by genetic, environmental, dietary and hormonal factors. The role of prolactin (PRL) in mammary tumorigenesis in rodents is well accepted but its involvement in human breast cancer has been controversial. PRL is a pleiotropic hormone that is produced by pituitary and non-pituitary sites under dissimilar regulatory mechanisms. Recent studies revealed expression of PRL and its receptor (PRL-R) in the human breast but the cell type that produces PRL, the regulation of its expression and whether local PRL promotes breast cancer growth in vivo have not been resolved. Preliminary Results: We found de-novo synthesis and time-dependent increases in PRL expression and release from human breast explants. PRL production was significantly higher in adipose than in glandular or malignant tissues. PRL expression was induced early during differentiation of breast preadipocytes whereas the expression of its receptor (PRLR) increased progressively, paralleling the rise in ieptin. Both PRL and the PRL-R were also expressed in two human liposarcoma cell lines. To test the concept that locally-produced PRL promotes tumor growth, the PRL transcript was stably transfected into breast cancer cells. Clones overexpressing PRL grew faster than controls in vitro and formed faster growing tumors in nude mice. Tumors derived from the PRL-overexpressing clones had increased levels of the PRL-R and the anti-apoptotic agent Bcl-2. Our main premise is that breast adipose tissue is the major source of local PRL which acts as a cytokine to promote tumor growth. To gain more understanding of breast adipose PRL, we will first determine the biochemical properties and regulation of PRL and its receptor in breast adipose tissue and preadipocytes. We will then generate cancer cells with a tetracycline-inducible PRL expression and analyze the mechanism by which paracrine/autocrine PRL stimulates tumor growth in vitro and in vivo. Hypothesis: Adipose PRL production is normally suppressed by tonic inhibition, but tumors produce PRL stimulatory factors that overcome this inhibition. The rise in local PRL activates mitogenic and anti-apoptotic signaling that stimulate tumor growth. Specific Aim 1 will characterize PRL and its receptor in breast adipose tissue. \ Specific Aim 2 will define the control of PRL expression in breast preadipocytes and liposarcoma cells. Specific Aim 3 will examine the effects of inducible PRL expression on tumor growth and gene expression. Long Term Goal: To establish PRL as a mitogen/anti-apoptotic factor in breast cancer, serving as the foundation for the design of future breast cancer therapy.